Project Summary Ribosome assembly is an essential process in all domains of life. In eukaryotes, in excess of 200 non- ribosomal factors are involved in this poorly understood process. Defects in human ribosome assembly are associated with ribosomopathies, a set of diseases, which result in high incidences of cancer. Our understanding of eukaryotic ribosome assembly is currently limited by an inability to stop this process at particular stages. We have recently developed a new technique to study early events of eukaryotic ribosome assembly as a function of transcription. This technique can recapitulate and arrest early eukaryotic ribosome assembly intermediates, thereby facilitating the analysis of eukaryotic ribosome biogenesis at an unprecedented level. In conjunction with other synergistic genetic, biochemical and structural biology methods, this technique will be used to elucidate hallmarks of early eukaryotic ribosome assembly, which previously represented intractable problems. Insights obtained from these studies will not only set new paradigms for eukaryotic ribosome assembly, but also have the potential to further our understanding of this process for the development of treatments.